LOCUS       sequence-253057-        6500 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of EGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6500)
  AUTHORS   Vo BT, Wolf E, Kawauchi D, Gebhardt A, Rehg JE, Finkelstein D, Walz 
            S, Murphy BL, Youn YH, Han YG, Eilers M, Roussel MF
  TITLE     The Interaction of Myc with Miz1 Defines Medulloblastoma Subgroup 
            Identity.
  JOURNAL   Cancer Cell. 2016 Jan 11;29(1):5-16. doi: 
            10.1016/j.ccell.2015.12.003.
  PUBMED    26766587
REFERENCE   2  (bases 1 to 6500)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.ccell.2015.12"; journalName: "Cancer Cell"; date: 
            "2016-01-11- 11"; volume: "29"; issue: "1"; pages: "5-16"
COMMENT     Sequence Label: pCDNA3-GFP
FEATURES             Location/Qualifiers
     source          1..6500
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    895..975
                     /label=MCS
                     /note="multiple cloning site"
     regulatory      977..986
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(983..985,986..988,989..1699,1700..1705,1706..1828)
                     /codon_start=1
                     /product="EGFP destabilized by fusion to residues 422-461 
                     of mouse ornithine decarboxylase, giving an in vivo 
                     half-life of ~2 hours"
                     /label=d2EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYKKLSHGFPPEVEEQDDGTLPMSCAQESGMDRHPAACASARINV
                     "
     primer_bind     complement(1028..1049)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1289..1308)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1636..1657
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1950..2071
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2091..2108)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2097..2321
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2367..2795
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2454..2473)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2664..2685
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2804..2824)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2809..3138
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2989..3124
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3051..3070
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3205..3999
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3259..3278)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3869..3888
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4173..4294
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4343..4359)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4343..4359)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4356..4378)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4367..4383
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4391..4397,4398..4415,4416..4421))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4436..4457
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4574..4591)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4745..5333)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4825..4844)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5504..6295,6296..6364))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6127..6146
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6365..6469)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gctaccggac tcagatctcg agctcaagct tcgaattctg cagtcgacgg taccgcgggc
      961 ccgggatcca ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt
     1021 gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga
     1081 gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa
     1141 gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag
     1201 ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta
     1261 cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt
     1321 gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga
     1381 ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat
     1441 catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga
     1501 ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc
     1561 cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa
     1621 cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg
     1681 catggacgag ctgtacaaga agcttagcca tggcttcccg ccggaggtgg aggagcagga
     1741 tgatggcacg ctgcccatgt cttgtgccca ggagagcggg atggaccgtc accctgcagc
     1801 ctgtgcttct gctaggatca atgtgtaggc ggccgcgact ctagatcata atcagccata
     1861 ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga
     1921 aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca
     1981 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     2041 gtggtttgtc caaactcatc aatgtatctt aagtttaaac cgctgatcag cctcgactgt
     2101 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     2161 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     2221 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     2281 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     2341 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     2401 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     2461 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     2521 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     2581 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     2641 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     2701 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     2761 aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta
     2821 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     2881 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2941 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     3001 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     3061 gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     3121 ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa
     3181 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     3241 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     3301 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     3361 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     3421 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     3481 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     3541 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     3601 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     3661 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     3721 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     3781 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     3841 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     3901 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     3961 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
     4021 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
     4081 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     4141 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     4201 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     4261 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
     4321 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     4381 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     4441 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     4501 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     4561 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     4621 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     4681 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     4741 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     4801 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     4861 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     4921 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     4981 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     5041 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     5101 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     5161 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     5221 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     5281 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     5341 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     5401 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     5461 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     5521 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     5581 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     5641 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     5701 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     5761 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     5821 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     5881 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     5941 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     6001 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     6061 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     6121 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     6181 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     6241 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     6301 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     6361 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     6421 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     6481 gaaaagtgcc acctgacgtc
//
